Cancer Therapy: Preclinical Novel Oligoamine Analogues Inhibit Lysine-Specific Demethylase 1 and Induce Reexpression of Epigenetically Silenced Genes

Purpose: Abnormal DNA CpG island hypermethylation and transcriptionally repressive histone modifications are associated with the aberrant silencing of tumor suppressor genes. Lysine methylation is a dynamic, enzymatically controlled process. Lysinespecific demethylase 1 (LSD1) has recently been identified as a histone lysine demethylase. LSD1 specifically catalyzes demethylation of mono– and dimethyl–lysine 4 of histone 3 (H3K4), key positive chromatin marks associated with transcriptional activation. We hypothesized that a novel class of oligoamine analogues would effectively inhibit LSD1 and thus cause the reexpression of aberrantly silenced genes. Experimental Design: Human colorectal cancer cells were treated with the oligoamines and changes in monoand dimethyl-H3K4 and other chromatin marks were monitored. In addition, treated cells were evaluated for the reexpression of the aberrantly silenced secreted frizzled-related proteins (SFRP) Wnt signaling pathway antagonist genes. Finally, the effects of the LSD1 inhibitors were evaluated in an in vivo xenograft model. Results: Treatment of HCT116 human colon adenocarcinoma cells in vitro resulted in increased H3K4 methylation and reexpression of silenced SFRP genes. This reexpression is also accompanied by a decrease in H3K9me2 repressive mark. Importantly, cotreatment with low doses of oligoamines and a DNA methyltransferase inhibitor highly induces the reexpression of the aberrantly silenced SFRP2 gene and results in significant inhibition of the growth of established tumors in a human colon tumor model in vivo. Conclusions: The use of LSD1-inhibiting oligoamine analogues in combination with DNA methyltransferase inhibitors represents a highly promising and novel approach for epigenetic therapy of cancer. (Clin Cancer Res 2009;15(23):7217–28) Epigenetics refers to heritable changes in gene expression patterns that are not regulated by changes in the primary DNA sequence. In cancer, epigenetic silencing of gene expression, including of tumor suppressor genes, is a common occurrence (1) that is associated with abnormal DNA methylation patterns and changes in covalent histone modifications (2). The amino-terminal tails of histones are subject to several posttranslational modifications, including acetylation, phosphorylation, and methylation, which are closely tied to transcriptional regulation, DNA replication, and DNA repair (2). As shown by the regulation of histone acetylation by histone acetyltransferases and histone deacetylases (HDAC), the addition and removal of these posttranslational modifications is a dynamic process. A similar dynamic regulation occurs for histone methylation with histone methyltransferases for the addition of methyl groups, and we recently discovered families of enzymes for specific histone demethylation. The first of these demethylating enzymes identified was the lysine-specific demethylase (LSD1/KDM1; ref. 3), a flavin adenine dinucleotide (FAD)-dependent amine oxidase, which interacts directly with CoREST and HDAC1/2 proteins, forming a module found in several multiprotein corepressor complexes and is known to act on intact chromatin as part of these complexes (3, 4). LSD1 demethylates H3K4me2/me1 through an oxidative reaction that leads to the reduction of the protein-bound FAD cofactor and the production of H2O2 and formaldehyde. More recently, a number of Jumonji (JmjC) domain–containing histone demethylases have been identified and shown to play important roles in concert with other histone-modifying enzymes related to the control of transcriptional regulation, cellular differentiation, and animal development (5, 6). Authors' Affiliations: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, The Johns Hopkins University School of Medicine, Baltimore, Maryland; Progen Pharmaceuticals, Redwood City, California; and Department of Pharmaceutical Sciences, Wayne State University, Detroit, Michigan Received 5/21/09; revised 8/7/09; accepted 8/24/09; published OnlineFirst 11/24/09. Grant support: NIH grants CA 51085 and CA 98454, Susan G. Komen for the Cure Foundation KG 088923, and Samuel Waxman Cancer Research Foundation. Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). Requests for reprints: Robert A. Casero, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Room 551, Bunting-Blaustein Cancer Research Building, 1650 Orleans Street, Baltimore, MD 21231. Phone: 410-955-8580; Fax: 410-614-9884; E-mail: [email protected]. F 2009 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-09-1293 7217 Clin Cancer Res 2009;15(23) December 1, 2009 www.aacrjournals.org Research. on April 12, 2017. © 2009 American Association for Cancer clincancerres.aacrjournals.org Downloaded from Published OnlineFirst November 24, 2009; DOI: 10.1158/1078-0432.CCR-09-1293